The retinoblastoma tumor susceptibility gene RB, located at 13q14.2, was originally identified in the childhood cancer retinoblastoma. It was one of the first tumor suppressor genes to be characterized and its product (pRB) is functionally inactivated in several human cancers. Among the functions of pRB is its ability to bind E2Fs and to inhibit E2F1-mediated transcription. In many tumor cells, the inactivation of pRB deregulates E2F activity and increases in cell proliferation. Uncontrolled E2F1 drives cell proliferation but it also induces apoptosis in response to specific cues such as DNA damage. We found that human colorectal cancer cells retain an intact RB gene and appear not to follow this paradigm. Another well common oncogenic pathway is the Wnt/ 2 - -catenin pathway. We have identified a functionally antagonistic relationship exists between these two pathways. Specifically, E2F1 abrogates 2 - -catenin activity generating selection for copy gains and overexpression of pRB and CDK8 in human colorectal cancer. Consequently, pRB and CDK8 act to suppress the inhibitory effect of E2F1 on 2 - -catenin transcriptional activity, allowing continued proliferation of colorectal cancer cells. I hypothesize that some human cancer cells are pRB-dependent and that the intact RB gene contributes to oncogenesis through increased 2 - -catenin transcriptional activity. In support of this, preliminary data shows that some human colorectal cancer cells (but not others) require pRB for rapid proliferation. I propose to carry out a screen of 43 human colorectal cancer cell lines to determine the requirement for pRB in colorectal cancer (Aim I). Uncontrolled proliferation occurs as a result of deregulated 2- -catenin transcriptional activity and with our findings that ectopic E2F1 expression abrogates this effect and CDK8 expression rescues it, I will also determine the consequences of misregulation of CDK8, 2 - -catenin, and E2F1. I will also look into the functional relationship between E2F1 and 2 - -catenin (Aim II). E2F1 inhibits 2 - -catenin activity, but the mechanisms underlying this control are unclear. I propose to determine the level at which E2F1 inhibits 2 - -catenin activity and will test the potential involvement of specific E2F1 target genes in the inhibition of 2 - -catenin activity. In addition to colorectal cancer cells, we found through database searches of human tumor tissues and cancer cell lines, that RB is upregulation and chromosome 13 copy gains and losses in human cancers other than colorectal cancer. I propose to screen through these tumor tissue and cancer cell lines to determine whether any other human cancers are pRB-dependent (Aim III). If pRB dependency is found among human cancers (other than colorectal cancer), then mechanistic pathways can be dissected in a similar manner as in Aim II. Together, these experiments will characterize an unexpected and novel activity for the RB tumor suppressor.